Assessment of Hearing Loss in Two-Year Follow-up Study of Neonates with Congenital Cytomegalovirus Infection

Objectives Congenital Cytomegalovirus (cCMV) infection constitutes the main cause of sensory neural hearing loss (SNHL) worldwide. The rate of Cytomegalovirus (CMV)-induced SNHL is not well documented in developing countries, such as Iran. Therefore, this prospective follow-up study aimed to evaluate this rate among neonates with cCMV infection in Iran. Materials & Methods Neonates with cCMV infection admitted to neonatal intensive care units and neonates with CMV infection identified in two other prospective screening studies in Tehran, Iran, were enrolled in this study. Audiological assessments, including otoacoustic emission and auditory brainstem response tests, were performed for all the cases. Antiviral therapy was administered for the newborns in case of having severe symptoms. Results A total of 22 neonates with cCMV infection were entered into the study, of whom 8 and 14 subjects had symptomatic and asymptomatic cCMV infection, respectively. In total, 3 of 22 newborns had SNHL (13.6%; 95% CI: 2.8-39.8), 2 of 8 cases with symptomatic cCMV infection (25.0%; 95% CI: 3-90) and 1 of 14 cases with asymptomatic cCMV infection (7.1%; 95% CI: 0.1-39). No association was observed between SNHL and CMV-related risk factors in newborns. Conclusion The findings of this study revealed that the rate of cCMV-induced SNHL is high among neonates born in Tehran. The severe sequelae of cCMV infection indicate the need for screening for CMV infection at birth to reduce the risk of CMV complications and the financial load of treatment imposed on healthcare and treatment systems in Iran.


Introduction
Congenital Cytomegalovirus(cCMV) is the leading cause of congenital anomalies and neurological damage in children worldwide (1). The annual incidence of cCMV infection is estimated within the range of 2-22 per 1000 live births in the world (2). It is important to note that due to postnatal exposure to Cytomegalovirus (CMV), newborns might shed the virus in their urine or saliva after 21 days of birth. Therefore, the diagnosis of cCMV infection should be made within the first 3weeks of the neonate's life (3).
Generally, more than 90% of newborns with cCMV infection are asymptomatic at birth, and of the remaining 10% infected neonates, numerous cases go unidentified due to the lack of specific symptoms or signs of CMV infection (4). Unfortunately, the complications of cCMV infection with continuing risk through adult life encompass lifelong disabilities, such as sensory neural hearing loss (SNHL) (i.e., hearing loss [HL] due to the auditory nerve and/or cochlear damage) and neurological impairment (5). It is estimated that about 25% of all cases with HL (in children up to 5years of age) are related to cCMV infection, less than one-third of which are recognized in newborn hearing screening due to the late onset of this sequela (6).
Moreover, it is estimated that approximately 9.3-17% of neonates with cCMV infection will develop SNHL during childhood (7,8). It is important to note that this viral infection is known as the leading non-genetic cause of SNHL worldwide.
Given that, universal CMV screening and targeted CMV screening of neonates who failed in neonatal hearing screening tests are two approaches that gained more attention (9

Materials & Methods
In this prospective follow-up study, neonates with Total DNA was isolated from the DBS samples in triplicate using heat shock assay according to a modified protocol (14). Briefly, the DBS samples were soaked in Minimum Essential Medium (Sigma-Aldrich Co., USA) and incubated at 4°C overnight and then heated by a thermocycler with a program at 55°C for 60 minutes, 100°C for 7minutes, and0°C for 2minutes (Eppendorf, Germany). Then, the DBS samples were

Results
Of 23 eligible newborns with confirmed cCMV infection, 1caserelocated to another city and was excluded from the study. Moreover, 9 and 13 of 22 neonates were female and male, respectively.

Clinical Findings
In this study, 14 of 22 infected neonates presented jaundice after birth, of whom 10 subjects did not require phototherapy; however, 4 subjects had severe jaundice and underwent phototherapy.
The peripheral blood smear of seven neonates revealed few erythroblasts and pancytopenia.
Germinal matrix hemorrhage was observed in the brain ultrasound of one infected newborn. In two neonates, the abdominal ultrasound scan revealed hepatomegaly and splenomegaly; nevertheless, the kidney size was normal. Respiratory distress syndrome was observed in one infected neonate; nonetheless, the chest X-ray was normal. The brain CT scan in one newborn revealed mild hydrocephaly with no signs of calcification.
When this neonate was treated with ganciclovir, a significant reduction was observed in spleen size; however, no increase was observed in head circumference. In ophthalmologic assessment, retinal hemorrhage without the inflammation of systemic deficits was detected in one neonate.
Additionally, chorioretinitis was reported in one infected newborn after 9 months of birth.

Follow-up
All infected neonates completed a follow-up study

Maternal Factors
The results of CMV serologic status during the first trimester of pregnancy were available for 17 of 22 mothers with CMV infected neonates, of whom11 subjects were previously infected with CMV and classified as women with non primary CMV infection (based on the presence of CMV immunoglobulin G antibody in their serum samples) and 6 subjects were infected newly with CMV and classified as mothers with primary CMV infection. Table 2 shows the data on the association between HL in newborns and the type of CMV infection in their mothers. Additionally, the level of CMV awareness among neonates' parents was very low, and none of them had heard about CMV before this project.

In Conclusion
The findings of this study demonstrate that cCMV